Construction Of PH And Redox Responsive Polyzwitterion Nano Carriers For Tumor Targeting Drug Delivery

Recently,nano drug delivery systems(NDDS)with their nanoscale size,good stability and drug loading capacity,promoted the drug accumulation in tumor site with reduced side effects,which have emerged as indispensable platforms for cancer theranostics.However,most clinically used nano drugs do not gain significantly improved treatment effect.Gaps between the potential of NDDS and actual therapeutic outcomes indicate the discrepancy between carrier design and human body physiological environment.Before a drug can take action inside tumor cell,it must complete an extensive journey,which begins with the injection of the drug carrier into the bloodstream and continues through stages of circulation,extravasation,accumulation and distribution,endocytosis,endosomal escape,intracellular localization and action.Most nanoparticles(NPs)specialize in one or two of these stages,but each is critical to the overall success of therapy.For example,body fluid dilution effect in the process of the systemic circulation,clearance of reticuloendothelial system(RES),uncertainty of the enhanced permeability and retention effect(EPR)in some types of tumor and inhomogeneity of vascular permeability,low drug bioavailability results from inefficient drug release leading to decreased drug efficacy and increased possibility of multidrug resistance(MDR)etc.Therefore,to improve the tumor targetting drug delivery efficiency,the structure and composition of nano carriers should be rantionally designed according to the physiologic barriers and characteristics of tumor microenvironment.Based on the above research background,pH and redox responsive zwitterion polymeric NDDS were designed on basis of physiological barriers and the characterization of tumor microenvironment.In order to improve therapeutic effect,the structure-activity relationship was investigated for synergistically exerting functions of drug loading,prolonged systemic circulation,tumor cell targeted drug release.In summary,this dissertation includes four parts as follows:(1)Amphiphilic RGD-PCB-b-PDPA(RGD-PCD)copolymers were prepared by continuous reversible addition fragmentation chain transfer(RAFT)polymerization using CB-tBu and DPA as monomers.Doxorubicine(DOX)was encapsulated as model medicine(RGD-PCD/DOX NPs).Research results of pH sensitivity of nanoparticle assembly behavior and drug release profile indicated that RGD-PCD/DOX NPs were stable at pH 7.4 with low drug release rate;when pH was lower than 6.5,protonation of the PDPA blocks led to the disassembly of NPs with accelerated drug release.In addition,results of fluorescence microscopy and flow cytometry analysis demonstrated improved internalization of RGD-PCD/DOX NPs in HepG2 cells via integrin-mediated endocytosis with rapid DOX release intracellularly.Furthermore,the RGD-PCD/DOX NPs exhibited prolonged systemic circulation and enhanced tumor accumulation in tumor-bearing nude mice in comparation with free DOX and non-targeting PCD/DOX NPs.(2)Reduction-responsive amphiphilic block copolymers,incorporating disulfide linkage between hydrophobic and hydrophilic blocks in main backbone,proved to be powerful carriers to facilitate intracellular drug delivery,owing to the concentration gradient of GSH between body fluids and the cytosol of tumor cells.In order to prepare cRGD-PCSSL diblock copolymers without using thiol exchange reaction,novel bioreducible double-head agent DDMAT-SS-OH was firstly synthesized,which connected RAFT agent and ring opening polymerization(ROP)initiator through disulfide bond.Drug encapsulated zwitterionic NPs were constructed by one-step self-assembly with DOX(termed as cRGD-PCSSL/DOX NPs).The reduction-responsiveness of cRGD-PCSSL/DOX NPs was measured by DLS and TEM under conditions mimicking in vivo situation.Results demonstrated that cRGD-PCSSL/DOX NPs could maintain stable particle size and morphology in the presence of GSH(20 μM),while lost stability gradually with the incubation of GSH(10 mM),which led to shedding of PCB shells through disulfide bond breaking.The results of pharmacokinetics and tissue distribution study showed that low protein adsorption and excellent serum stability properties of cRGD-PCSSL/DOX NPs translated into prolonged systemic circulation and enhanced tumor accumulation.Furthermore,intravenous injected cRGD-PCSSL/DOX NPs in tumor-bearing balb/c nude mice exhibited significantly higher antitumor efficiency and lower cardiotoxicity compared to free DOX.(3)Triblock RGD-PCSSD copolymers were prepared to construct nanocarriers,which cooperatively combined RGD ligand for active tumor targeting and receptor-mediated endocytosis,ultra low fouling zwitterionic PCB corona for prolonged systemic circulation,disulfide-crosslinked shell for enhanced circulation stability and drug loading stability,highly pH-sensitive PDPA core for efficient drug encapsulation and responsive release.Study of cooperativity between pH sensitive core and redox-responsive shell showed that crosslinked shell had inhibiting effect on the protonation of PDPA core of PCSSD SCNPs,which increased with the increase of crosslinking density.In other words,the balanced point between the core expansion force induced by protonation and the shell crosslinking caused restrictive effect could be teamed as pKa* value.Furthermore,the cooperativity and dual sensitivity properties of well-defined SCNPs could efficiently restrain drug leakage during systemic circulation and in the matrix of tumor tissues,while implement “spatiotemporally pinpointed” drug release intracellularly.(4)On basis of the above studies,redox responsive self-immolation polymers were designed to prepare “lock-key” combined nanocarrier.Firstly,carbonic ester monomer TMC-DS was synthesized.However,no polymer was obtained acoording to reported polymerization methods.In order to analyse the reason,TMC-DS and TMC-EA monomers were synthesized.MTC-DS monomer could be polymerized using TBD as the catalyst instead of TMC-EA,indicating that the disulfide bond connected with the six-membered ring of carbonic ester affect the polymerization process,but the mechanism need to be further investigated.In addition,to continue the research on the relationship between polymer self-immolation property and assembly behavior of coresponding NPs,TMC-N-DS monomer was synthesized to prepare SIPs with two step self-immolation property.After incubation of prepared NPs with GSH(10 mM)for 12 h,the disulfide bonds were reduced to sulfydryl,however,no disassembly was detected.Therefore,SIPs should be further optimized and the effect factors of self-immolation rate should be investigated.In summary,a series of p H and redox responsive zwitterion polymeric NDDS were prepared and systemically evaluated in vitro and in vivo,which could provide potential carriers for antitumor drugs and reference for stimuli responsive nanocarrier design.